Lichenoid areas may arise in early stages of proliferative verrucous leukoplakia: A long‐term study of 34 patients

Abstract Background Proliferative verrucous leukoplakia is considered an uncommon oral potentially malignant disorder with a high malignant transformation rate. The objective of this paper was to define its cancer incidence and related risk factors. Methods A retrospective audit of 34 patients diagnosed with proliferative verrucous leukoplakia from a university‐based unit, during the period from 1995 to 2019 was performed. The mean number of visits was 23 ± 18.6. The follow‐up was divided into four‐time intervals to evaluate the clinical presentation, number of lesions, dysplasia grade, and malignant transformation rate. Results The majority of patients were females 29 (85.3%), with verrucous component (77.8%), with a gingival presentation (31.8%), and with a preceding lichenoid area (44.1%). Eleven patients (32.4%) were affected by oral cancer during the follow‐up, developing a total of 15 carcinomas. The mean age of malignant transformation was 67.2 ± 12.9 years, particularly 8 ± 8.5 from the onset of the lesions. Warty forms presented a higher mean estimate for malignant transformation (15.2 years, 95% confidence interval 4.4–26 years) than nodular forms (1.9 years, 95% confidence interval 1.9–1.9) (p = 0.019). Patients with an initial proliferative verrucous leukoplakia diagnosis suffered a higher risk of malignancy, particularly 15.55 times (95% confidence interval 1.69–143.17; p = 0.015) than those who did present a preceding area with lichenoid morphology. Conclusion Proliferative verrucous leukoplakia presented a high malignant transformation rate and sometimes displayed preceding oral lichenoid areas in early stages. Further studies are needed to understand the impact of these lichenoid areas in proliferative verrucous leukoplakia progression.


| INTRODUCTION
Oral leukoplakia (OL) is considered the most prevalent and significant oral potentially malignant disorders (OPMDs) worldwide. OL is defined as "a predominantly white plaque of questionable risk having excluded (other) known diseases or disorders that carry no increased risk for cancer." 1,2 The last World Health Organization (WHO) Collaborating Centre for Oral Cancer position paper regarding OPMDs nomenclature found no logical reason to change this classical definition. Nonetheless, this working group decided to define separately classical OL from a particular form of this disorder, namely proliferative verrucous leukoplakia (PVL). 3 The term PVL was first coined more than 35 years ago by Hansen et al. 4 Since then, individual reports, small case series, and reviews have been published regarding this enigmatic form of OL. 5 Despite these continuous efforts, PVL diagnosis criteria remained unresolved for decades. Indeed, even the aetiopathogenesis and associated risk factors of these OPMDs remain unclear. In order to unify reporting guidelines, this latest WHO working group ended up defining PVL as a "progressive, persistent, and irreversible disorder characterized by the presence of multiple leukoplakias that frequently become warty." 3 It is worth mentioning that PVL does not carry a histopathologic connotation but subtle pathologic features, such as the presence of lichenoid chronic inflammation, dense/wavy hyperkeratosis, acanthosis, papillomatous squamous proliferation, and variable degrees of dysplasia. 6 Moreover, this clinical entity is well known for its marked tendency to recur. In this sense, a recent meta-analysis yielded a pooled recurrence rate of 67.2%. 7 The most important complication of PVL is the development of oral cancer. 8 This fact carries relevant implications for the management of these patients, especially in regard to the surveillance programs that they must receive, given that these disorders are likely to accompany them during all their lives. 2 The malignant transformation (MT) rate of this OPMD is among the highest of its spectrum ranging from 43.8% to 65.8%. 9,10 In addition, several authors have considered that field cancerization exerts a relevant effect on this disorder leading frequently to multiple primary tumors in these patients. 11 Taking together, the crucial point for PVL diagnosis and management is the keen observation of its gradual topographical, and histopathological changes. 12 In the authors' experience, we consider the proliferative or multifocal nature of the disease as the main cornerstone for diagnosis. 9 Other researchers consider the presence of verrucous features as a main diagnostic criteria, or even the same authors consider all gingival leukoplakias regardless of size to be PVLs. 13 These factors for establishing a PVL diagnosis case may be over-restrictive and even confusing, implying a potential underdiagnosis and misestimation of the true malignant transformation rate of this uncommon but relevant OPMD.
Prompted by discussed literature, we undertook a retrospective study of a carefully documented case series to elucidate any early features or risk factors noted in clinical or histopathological records. The objective of the study was twofold: (i) to describe the clinical and histopathological features of the lesions during the follow-up; and (ii) to investigate the cancer incidence in patients with this OPMD and its associated risk factors. Initially, all patients were biopsied to establish diagnoses. Particularly, we used an incisional technique involving the removal of a representative portion of the target white patch but also a part of adjacent healthy tissue. Due to the multifocal nature of PVL, different samples in each intervention were obtained, placing each of them in a separate and adequately identified container for further histopathological analysis. During follow-up, we opt to perform biopsies upon the detection of an oral anomaly (e.g., new ulceration, tumor, or tissue growth in mucosa). Selected regions were chosen on the basis of a conventional oral examination performed by two trained specialists in oral medicine (ABC and PGV) using 2.5/3 magnification loupes with white LED headlight, particularly BLD-3 (Neitz Instruments Co., Ltd.). The samples were fixed in formaldehyde and were subjected to a histopathological evaluation. Pathology reporting on oral epithelial dysplasia for this case-series was presented according to binary and the WHO 2005 histologic grading systems. 15,16 This analysis was performed by senior oral pathologists calibrated to each other, as previously reported. 17 All patients were informed that they had an OPMD, stressing the high frequency of malignant transformation and allowing them sufficient time to raise all their doubts or questions. Patients were advised of the need for regular follow-ups over their lifetime, given the unpredictability of oral cancer development. Follow-ups were programmed 2 months after initial diagnosis. Patients were instructed to contact the clinician between visits immediately upon the detection of an oral anomaly or suspicion also to clarify any doubt that might arise. All patients who missed a follow-up session were contacted by phone.
The mean number of visits was 23 ± 18.6. In order to tackle the problem of multiple periods of follow-up among patients, this period was divided into time intervals. Globally, follow-up was divided into four intervals segmented as follows: T0 the initial visit with a histopathological assessment, T1 1 /4 of the follow-up, T2 3 /4 of the followup, and TF the last recorded appointment.
In the event of a malignant transformation, it was collected at the closest corresponding interval. WHO malignant transformation definition (i.e., biopsy-proven PVL) was used in this audit, including only those cases of PVL that have had their diagnosis confirmed by both clinical examination/photographs and histological assessment prior to malignancy. Information on the age at diagnosis, gender, presence of lichenoid areas, unhealthy oral habits, systemic health status, location, number of lesions, date of diagnosis, and treatment for PVL such as cold knife excision, or laser therapy was collected from clinical charts.
A pre-tested questionnaire was posted to all participants who were on follow-up in order to avoid data attrition.

| Characteristics of study population and clinical presentation
A total of 34 patients were eligible for the study with a complete data set ( Figure 1). Table 1  Considering the four intervals of follow-up, gingiva was the most frequent location (31.8%), followed by the buccal mucosa (30.4%).
Regarding the number of affected areas, they are extensively displayed in Figure 2 as a heat-map. The larger the follow-up, the more pronounced number of affected regions and higher number of lesions.
Paradoxically, treatment modality did not exhibit an effect on these outcomes. The global proportion rate for recurrence was 23.5% with no significant differences between both arms of therapies. None of the participants died as a consequence of the disease at the end of the follow-up.

| Rate of malignant transformation and associated factors
The mean follow-up time from diagnosis was 5.7 ± 4.1 years with a range from 0.6 to 14.3 years. In this time frame, 11 patients (32.4%) were affected by oral cancer, developing a total of 15 carcinomas (three patients suffered two primary de novo malignancies). Among these oral cancers: nine were conventional squamous cell carcinomas, three were verrucous carcinomas, and three were in situ carcinomas.
The mean age of malignant transformation was 67.2 ± 12.9, particularly 8 ± 8.5 from the onset of the lesions.   We also verified the typical multifocal presentation of this disorder. The multifocal presentation of PVL and its risk of multiple primary oral carcinomas have been associated with the concept of field cancerization originated in 1953 from the histopathological observations of Slaughter and colleagues. 21 This concept was introduced to describe the occurrence of multifocal neoplastic lesions in the oral epithelium, particularly in the adjacent non-tumor epithelia of oral carcinomas.
Potentiating factors, such as smoking or alcohol intake, have not demonstrated any evident influence on either occurrence or progression in the present study. It is also worth mentioning that the consumption of these carcinogens was anecdotal in our study. This is consistent with previous case series addressing this issue. 2,10 The consumption of tobacco and alcohol does not seem to play an important role in the onset of this lesion This fact differs particularly from classical OL, for which tobacco is recognized as the main risk factor. 22 Borgna et al. 20 found no significant differences regarding smoking and alcohol use between 23 patients with PVL who progressed to malignancy and the 25 individuals with PVL without progression.
In agreement with studies of other predominantly Caucasian patient groups, PVL seemed to develop in late life and being more frequent in females without unhealthy habits. 23 In terms of gender, although females were more commonly affected by PVL than males, no association between gender and malignant transformation was noticed. A recent meta-analysis found that women have a statistically higher incidence of PVL, and an also increased risk for malignant transformation than males. 24 Moreover, the mean age of patients was higher than 60 years as in most published studies; however, no clear

CONFLICT OF INTEREST
The authors declare no conflict of interest.

DATA AVAILABILITY STATEMENT
The data that support the findings of this study are available from the corresponding author upon reasonable request.

INFORMED CONSENT
Written Informed consent was obtained for all patients participating in the study.